This work will involve investigations into the molecular mechanisms whereby insulin stimulates the sodium pump. The principal approach will involve studies on intact muscle cells and also upon the (Na, K ion)-ATPase isolated from the same cells. The validity of the kinetic analysis used to analyze the effect of insulin upon 22Na efflux (Moore, 1973) will be tested by using this technique to analyze the effect of catecholamines upon 22Na efflux and comparing the results to those using insulin. The stimulation of 22Na efflux by insulin into K-free Ringer containing ouabain will be investigated. Experiments will be conducted to determine if this represents exchange of intracellular Na ion for extracellular Na ion, Ca ions, or H ion. The response of hormone-induced uptake of 42K to ouabain will be determined and a comparison made between the effects of insulin and of catecholamines. The rate and mechanism of hyperpolarization produced by insulin and by catecholamines will be studied. The possibility that insulin increases the number of Na pump sites, as indicated by 3H-ouabain binding, will be investigated. The mechanism whereby insulin interaction with its receptors is transmitted to the Na pump will be studied. Three possible mechanisms for the mediation of this effect will be investigated: 1) Ca ions or Mg ions, 2) cyclic nucleotides (cyclic AMP or cyclic GMP), 3) mediation through a phase change within the membrane. The experiments will be conducted on frog skeletal muscle (sartorius) and upon rat skeletal muscle (soleus) with the muscle from one leg serving as a paired control for the other. Kinetic analysis of 22Na efflux will be performed by computer programs as outlined in Moore (1973). Membrane potentials will be measured by open tip glass microelectrodes and intracellular K ion activity will be followed with a glass microelectrode whose tip is filled by Corning 477317 K ion exchanger. Possible effects of insulin upon phase transitions in the plasma-membrane will be studied using Arrhenius plots of the effect of temperature upon the rate of the Na pump as measured by either 22Na efflux or by the rate of ATP hydrolysis catalyzed by the isolated (Na plus K ion)-ATPase.